Recombinant
RabMAb

Recombinant Anti-CRISPR-Cas9 antibody [EPR19633] (ab202657)

Overview

  • Product name

    Anti-CRISPR-Cas9 antibody [EPR19633]
    See all CRISPR-Cas9 primary antibodies
  • Description

    Rabbit monoclonal [EPR19633] to CRISPR-Cas9
  • Host species

    Rabbit
  • Tested applications

    Suitable for: WB, ICC/IF, IP, Flow Cytmore details
  • Species reactivity

    Reacts with: Other species
  • Immunogen

    Synthetic peptide within CRISPR-Cas9 aa 1000-1100. The exact sequence is proprietary. (Streptococcus thermophilus).
    Database link: G3ECR1

  • Positive control

    • WB: HEK-293 whole cell lysates transfected with CRISPR-Cas9 (G3ECR1, S.thermophilus, C–terminal aa801-1409) with GFP-Myc tag, CRISPR-Cas9 (G3ECR1) with Myc-His tag and CRISPR-Cas9 (Q03JI6, S.thermophilus (strain ATCC BAA-491 / LMD-9)) with Myc-His tag. ICC/IF: HeLa cells transfected with CRISPR-Cas9 (G3ECR1) with Myc-His tag. Flow Cyt: HEK-293 whole cell lysate transfected with CRISPR-Cas9 (G3ECR1) with Myc-His tag. IP: HEK-293 whole cell lysate transfected with CRISPR-Cas9 (G3ECR1) with Myc-His
  • General notes

     

     

    Our RabMAb® technology is a patented hybridoma-based technology for making rabbit monoclonal antibodies. For details on our patents, please refer to RabMab® patents.

    This product is a recombinant rabbit monoclonal antibody.

Properties

  • Form

    Liquid
  • Storage instructions

    Shipped at 4°C. Store at +4°C short term (1-2 weeks). Upon delivery aliquot. Store at -20°C long term. Avoid freeze / thaw cycle.
  • Storage buffer

    Preservative: 0.01% Sodium azide
    Constituents: 59% PBS, 40% Glycerol, 0.05% BSA
  • Concentration information loading...
  • Purity

    Protein A purified
  • Clonality

    Monoclonal
  • Clone number

    EPR19633
  • Isotype

    IgG

Applications

Our Abpromise guarantee covers the use of ab202657 in the following tested applications.

The application notes include recommended starting dilutions; optimal dilutions/concentrations should be determined by the end user.

Application Abreviews Notes
WB 1/20000. Detects a band of approximately 164 kDa (predicted molecular weight: 164 kDa).
ICC/IF 1/100.
IP 1/30.
Flow Cyt 1/60.

Target

  • Relevance

    [FUNCTION] CRISPR (clustered regularly interspaced short palindromic repeat) is an adaptive immune system that provides protection against mobile genetic elements (viruses, transposable elements and conjugative plasmids). CRISPR clusters contain spacers, sequences complementary to antecedent mobile elements, and target invading nucleic acids. CRISPR clusters are transcribed and processed into CRISPR RNA (crRNA) (Probable). In type II CRISPR systems correct processing of pre-crRNA requires a trans-encoded small RNA (tracrRNA), endogenous ribonuclease 3 (rnc) and this protein. The tracrRNA serves as a guide for ribonuclease 3-aided processing of pre-crRNA. Subsequently Cas9/crRNA/tracrRNA endonucleolytically cleaves linear or circular dsDNA target complementary to the spacer. The target strand not complementary to crRNA is first cut endonucleolytically, then trimmed by 3'-5' exonucleolytically. DNA-binding requires protein and both RNA species. Cas9 probably recognizes a short motif in the CRISPR repeat sequences (the PAM or protospacer adjacent motif) to help distinguish self versus nonself.
  • Alternative names

    • Cas9 antibody
    • CRISPR-associated endonuclease Cas9/Csn1 antibody
    • CRISPR-Cas9/Csn1 antibody
    • csn1 antibody
    • SpyCas9 antibody
    see all

Images

  • All lanes : Anti-CRISPR-Cas9 antibody [EPR19633] (ab202657) at 1/20000 dilution

    Lane 1 : Empty vector with GFP-Myc tag (vector control) transfected HEK-293 (Human epithelial cell line from embryonic kidney) whole cell lysate
    Lane 2 : HEK-293 (Human epithelial cell line from embryonic kidney) whole cell lysate transfected with CRISPR-Cas9 (Q99ZW2, Streptococcus pyogenes serotype M1) with GFP-Myc tag
    Lane 3 : HEK-293 (Human epithelial cell line from embryonic kidney) whole cell lysate transfected with CRISPR-Cas9 (G3ECR1, Streptococcus thermophilus, N–terminal aa1-800) with GFP-Myc tag
    Lane 4 : HEK-293 (Human epithelial cell line from embryonic kidney) whole cell lysate transfected with CRISPR-Cas9 (G3ECR1, Streptococcus thermophilus, C–terminal aa801-1409) with GFP-Myc tag
    Lane 5 : Empty vector with Myc-His tag (vector control) transfected HEK-293 (Human epithelial cell line from embryonic kidney) whole cell lysate
    Lane 6 : HEK-293 (Human epithelial cell line from embryonic kidney) whole cell lysate transfected with CRISPR-Cas9 (A1IQ68, Neisseria meningitidis serogroup A / serotype 4A (strain Z2491)) with Myc-His tag
    Lane 7 : HEK-293 (Human epithelial cell line from embryonic kidney) whole cell lysate transfected with CRISPR-Cas9 (G3ECR1, Streptococcus thermophilus) with Myc-His tag
    Lane 8 : HEK-293 (Human epithelial cell line from embryonic kidney) whole cell lysate transfected with CRISPR-Cas9 (Q03JI6, Streptococcus thermophilus (strain ATCC BAA-491 / LMD-9)) with Myc-His tag
    Lane 9 : HEK-293 (Human epithelial cell line from embryonic kidney) whole cell lysate transfected with CRISPR-Cas9 (J7RUA5, Staphylococcus aureus subsp. aureus) with Myc-His tag

    Lysates/proteins at 20 µg per lane.

    Secondary
    All lanes : Goat Anti-Rabbit IgG H&L (HRP) (ab97051) at 100000 cells

    Predicted band size: 164 kDa


    Exposure time: 5 seconds


    Blocking/Dilution buffer: 5% NFDM/TBST.

  • All lanes : Anti-CRISPR-Cas9 antibody [EPR19633] (ab202657) at 1/20000 dilution

    Lane 1 : HEK-293 (Human epithelial cell line from embryonic kidney) whole cell lysate transfected with CRISPR-Cas9 (G3ECR1, Streptococcus thermophilus, C–terminal aa801-1409) with GFP-Myc tag
    Lane 2 : HeLa (Human epithelial cell line from cervix adenocarcinoma) whole cell lysate
    Lane 3 : NIH/3T3 (Mouse embryonic fibroblast cell line) whole cell lysate
    Lane 4 : Rat embryo lysate

    Lysates/proteins at 20 µg per lane.

    Secondary
    All lanes : Goat Anti-Rabbit IgG H&L (HRP) (ab97051) at 1/100000 dilution

    Predicted band size: 164 kDa
    Observed band size: 86 kDa
    why is the actual band size different from the predicted?


    Exposure time: 5 seconds


    Blocking/Dilution buffer: 5% NFDM/TBST.

  • Immunofluorescent analysis of 4% paraformaldehyde-fixed, 0.1% Triton X-100 permeabilized HeLa (Human epithelial cell line from cervix adenocarcinoma) cells transfected with CRISPR-Cas9 (G3ECR1, Streptococcus thermophilus) with Myc-His tag or Empty vector, labeling CRISPR-Cas9 with ab202657 at 1/100 dilution, followed by Goat Anti-Rabbit IgG (Alexa Fluor® 488) (ab150079) secondary antibody at 1/1000 dilution.

    Confocal image showing Positive staining on HeLa cells transfected with CRISPR-Cas9(G3ECR1, Streptococcus thermophilus) with Myc-His tag.

    The nuclear counterstain is DAPI (blue).

    Tubulin is detected with Anti-alpha Tubulin antibody [DM1A] - Loading Control (ab7291) at 1/1000 dilution and Goat Anti-Mouse IgG H&L (Alexa Fluor® 594) preadsorbed (ab150120) at 1/1000 dilution (red).

    The negative controls are as follows:-

    -ve control 1: ab202657 at 1/100 dilution followed by ab150120  at 1/1000 dilution.

    -ve control 2: ab7291  at 1/1000 dilution followed by ab150077  at 1/1000 dilution.

  • Flow cytometric analysis of 4% paraformaldehyde-fixed HEK-293 (Human epithelial cell line from embryonic kidney) cells transfected with CRISPR-Cas9 (G3ECR1, Streptococcus thermophilus) with Myc-His tag labeling CRISPR-Cas9 with ab202657 at 1/60 dilution (red) compared with a Rabbit IgG,monoclonal [EPR25A]-Isotype control (ab172730)(black). Goat anti Rabbit IgG (Alexa Fluor® 488) at 1/2000 dilution was used as the secondary antibody.

  • CRISPR-Cas9 was immunoprecipitated from 0.35 mg of HEK-293 (Human epithelial cell line from embryonic kidney) whole cell lysate transfected with CRISPR-Cas9 (G3ECR1, Streptococcus thermophilus) with Myc-His tag with ab202657 at 1/30 dilution.

    Western blot was performed from the immunoprecipitate using ab202657 at 1/1000 dilution.

    VeriBlot for IP Detection Reagent (HRP) (ab131366), was used for detection at 1/10000 dilution.

    Lane 1: HEK-293 whole cell lysate transfected with CRISPR-Cas9 (G3ECR1, Streptococcus thermophilus) with Myc-His tag 10µg (Input).

    Lane 2: ab202657 IP in HEK-293 whole cell lysate transfected with CRISPR-Cas9 (G3ECR1, Streptococcus thermophilus) with Myc-His tag.

    Lane 3: Rabbit IgG,monoclonal [EPR25A]-Isotype Control (ab172730) instead of ab202657 in HEK-293 whole cell lysate transfected with CRISPR-Cas9 (G3ECR1, Streptococcus thermophilus) with Myc-His tag.

    Blocking and dilution buffer and concentration: 5% NFDM/TBST.

    Exposure time: 1 second.

References

ab202657 has not yet been referenced specifically in any publications.

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